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Absorption of protein digestion products: A review

D. M. MATTHEWS AND LEONARD LASTER

From the Department of Chemical Pathology, Westminster Medical School,London, and the Gastroenterology Unit, National Institute ofArthritis andMetabolic Diseases, National Institutes of Health, Bethesda, MD., U.S.A.

Each major phase in the evolution of our presentconcept of the absorption of proteins and theirdigestion products has been initiated by an improve-ment in analytical techniques or experimentalmethods.

Before 1900, it was thought that the final productsof protein digestion were proteoses and peptones,and it seemed likely that proteins were absorbed inthese forms. Then Cohnheim (1901) discovered anintestinal enzyme, 'erepsin', that catalysed thehydrolysis of these larger protein breakdownproducts to amino-acids. This was followed by thedemonstration that amino-acids were present in thesmall intestine during protein digestion (Kutscherand Seemann, 1901; Abderhalden, K6r6sy, andLondon, 1907), that animals could live on dietscontaining only hydrolysed protein (Loewi, 1902;Henderson and Dean, 1903) and could utilizeamino-acids given intravenously (Buglia, 1912), andthat there was a rise in the non-protein nitrogen ofthe blood during the absorption of protein, proteinhydrolysates, or amino-acids (Cathcart and Leathes,1906; Folin and Denis, 1912). These observationssuggested that proteins were absorbed from the gutand transported to the tissues as amino-acids, butsince the analytical methods then available were notsufficiently specific to show unequivocally thepresence of free amino-acids in the bloodstream, itwas possible to maintain that amino-acids might besynthesized into protein while passing through theintestinal wall and reach the tissues mainly in thisform (Abderhalden, 1912).

In 1912, Van Slyke and Meyer, using the recentlydeveloped nitrous acid method, showed that therewas a large rise in the amino-nitrogen of mesentericand peripheral blood during the absorption ofalanine and after feeding protein, and investigatedthe metabolic fate of absorbed amino-acids (VanSlyke and Meyer, 1912; 1913). Abel, Rowntree, andTurner (1914) obtained free amino-acids from theblood by vividialysis, and showed that they increasedafter a protein meal. This work laid the foundationfor the theory that protein was completely hydro-

lysed within the lumen of the gut, and that theconstituent amino-acids were absorbed into thebloodstream by simple diffusion (e.g., Verzar andMcDougall, 1936). This theory, though based onevidence that is now seen to be incomplete (e.g.,Dent and Schilling 1949; Fisher, 1954) seemedadequate to explain nearly all the older observationsand dominated the field for many years.

Further advances began in 1951, when it wasshown by means of stereochemically specific methodsthat the L-isomers of many amino-acids werepreferentially absorbed, which could not readily beexplained if absorption were by simple diffusion.The development of techniques for studying absorp-tion by isolated intestine in vitro made it possible toshow that L-amino acids were actively transportedby the small intestine, and since this time there hasbeen great progress in knowledge of the mechanismsof absorption of protein digestion products. Thispaper gives an account of modern views on thissubject and includes a brief description of theexperimental methods which have contributed mostto advances in the field.

TRANSPORT ACROSS THE INTESTINAL WALL

During absorption, a substance is removed from theintestinal lumen, transported across the intestinalwall and enters the blood or lymph. No singleexperimental technique will provide information onall these processes, nor are they all completelyelucidated in any instance. Much of the recent workon intestinal absorption has concerned the mode oftransport across the cells of the intestinal mucosa.The passage of substrates across the membranes ofthe mucosal cells is a special example of the generalprocess of membrane transport, a subject that hasbeen reviewed by Rosenberg (1954), Berliner (1959),and Christensen (1960). The structural features ofthe intestinal wall and their relation to the processesof absorption, and the nature of the mucosalbarrier, are discussed by Laster and Ingelfinger(1961) and by Wilson (1962; 1964).

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D. M. Matthews and Leonard Laster

Transport phenomena may be classified anddefined in many different ways, but it is convenientand simple to follow Crane (1960) in distinguishingtwo main modes of transport.

ACTIVE TRANSPORT Active transport is dependenton the expenditure of metabolic energy, and iscapable of moving substances across a membraneagainst a chemical or electrical gradient. Experi-mentally, the clearest evidence for active transportis the demonstration of transport against an electro-chemical gradient (i.e., 'uphill' transport). Confirm-atory evidence is provided by showing that thetransport is abolished by interfering with the meta-bolic energy supply, by such means as anoxia ormetabolic inhibitiors.

PASSIVE TRANSPORT Passive transport does notrequire the expenditure of metabolic energy, andoccurs in the same direction as the electrochemicalgradient that exists across the membrane. Thecharacteristics of passive transport are determinedby the nature of the molecules transported and theproperties of the membrane involved. The leastcomplex example of passive transport is 'simplediffusion' which occurs when a membrane is freelypermeable to the solute molecules. In simplediffusion, the rate of transport is directly propor-tional to the concentration difference across themembrane-an uncommon situation in biologicalsystems. There are comparatively few nutrients whichare now believed to be absorbed by simple diffusion.In a special form of passive transport known as'facilitated diffusion', the rate of transport is muchgreater than that which would be expected on thebasis of the known physical properties of thesystem. Certain features are common to bothfacilitated diffusion and active transport. Theseinclude selective transport of one substance ascompared with another of similar molecular size,competition for transport between structurallysimilar molecules, amd lack of proportionalitybetween transport rate and the concentrationdifference across the membrane. In both activetransport and facilitated diffusion, the transportrate tends to approach a limiting value as theconcentration of the transported substance isincreased and the transport mechanism becomessaturated. Active and passive transport are probablynot mutually exclusive, nor are they necessarilyentirely distinct processes. Thus it is quite probablethat a substance may be transported by bothprocesses simultaneously.The mechanisms responsible for active transport

and facilitated diffusion are poorly understood. Ahypothesis frequently used in discussions of active

transport is that of the 'shuttling carrier': that thecell membrane contains a carrier which can pick upa substance on one side and release it on the other,the system being 'driven' by metabolic energy. Asimilar hypothesis might explain facilitated diffusion,only in this case the system would not be driven bymetabolic energy. Other hypotheses of activetransport are described by Ussing (1957) andChristensen (1960; 1962).The view is sometimes expressed that active

transport mechanisms capable of moving solutesagainst concentration gradients are unlikely to beimportant in absorption from the intestine, since,under physiological conditions, solutes pass fromthe lumen of the gut to the blood in the direction of aconcentration gradient. Active transport, however,can probably greatly accelerate the movement ofsubstances which would diffuse only slowly throughthe intestinal wall. The diffusion of amino-acidsthrough non-living membranes is known to beunusually slow (Schmengler, 1933). It has also beensuggested that active transport may be important inlimiting back-diffusion from the blood into theintestinal lumen (Booth and Kanaghinis, 1963).

METHODS OF INVESTIGATINGINTESTINAL ABSORPTION

METHODS in vitro Much of the recent work on theabsorption of amino-acids and peptides has beencarried out using isolated intestine in vitro. Methodsin vitro are less 'unphysiological' than would appearat first sight (Smyth, 1961a), and provide infor-mation difficult or impossible to obtain in any otherway.

Early preparations of intestine in vitro did notgive very satisfactory results, probably owing toinadequate oxygenation of the mucosa. The firstsatisfactory preparation in vitro was that of Fisherand Parsons (1949), in which the lumen of a lengthof rat small intestine was perfused with oxygenatedsaline while the serosal surface was bathed in thesame solution. A simplified apparatus was devisedby Wiseman (1953) and used to demonstrate theactive transport of amino-acids; Darlington andQuastel (1953) described an apparatus based onsimilar principles.The simplest and most widely used method in

vitro is that using everted sacs of small intestine(Wilson and Wiseman, 1954). A length of smallintestine from a small animal, usually a rat orhamster, is everted with a glass rod so that themucosal surface is on the outside. Sacs are madefrom the everted intestine and filled with Krebs-Ringer saline, or other appropriate solution(usually 0 5-1 ml.); this is termed the 'serosal fluid'.

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Absorption ofprotein digestion products: A review

25ml. FLASK - - OXYGEN

SEROSAL MUCOSALFLUID D FLUID

_~-GUT SA

FIG. 1. A 'simple sac' of everted intestine.

Each sac is placed in a flask containing saline(usually 5-10 ml.); this is termed the 'mucosal fluid',and incubated under oxygen with gentle shaking(Fig. 1). The preparation remains active for at leastone hour at 37°C. Eversion of the sac, with slightdistension, stretches the mucosa and improves itsoxygenation; while the small volume of the fluidwithin the sac causes transport of any solute fromthe mucosal to the serosal side to produce a relativelylarge increase in concentration in the serosal fluid.In investigation of active transport, the experimentis usually begun with equal concentrations of thetest substance in mucosal and serosal fluids. If atthe end of the experiment, the concentration in theserosal fluid has risen and that in the mucosal fluidhas fallen, it can usually be taken that transportagainst a concentration gradient has occurred (Fig. 2).

a 0 -J c c J a

w LL LL ~~~~~~~~~~~U. UA.

GO. s a <<t(Ae<sehd0 0 0cenr n U U in

acosth walit h eoa lud ota hia

IL0z0

C-

wUz0U

INITIAL STATE FINAL STATE

FIG. 2. Transport against a concentration gradient. Atthe start of the experiment, the mucosal and serosal con-centrations of the substance transported, e.g., an amino-acid or glucose, are equal. At the end of the experiment,the substance has been accumulated to a high concentra-tion in the gut wall and much of it has been transportedacross the wall into the serosal fluid, so that the finalconcentration in the serosal fluid is higher than that in themucosal fluid.

This is, however, only true if the serosal volume hasincreased (as it usually does, owing to water trans-port) or at least remained constant; in other words,if the increase in concentration of the solute on theserosal side cannot be accounted for by net watermovement in the opposite direction, with retentionof solute in the serosal fluid. Transport of solutesout of the mucosal fluid and into the serosal fluidcan be measured from the changes in concentrationin these fluids, and from their initial and finalvolumes.

In the technique of Crane and Wilson (1958), theeverted sac is cannulated at one end (Fig. 3). Withthis preparation, the time course of transport intothe serosal fluid can be followed by serial sampling,and it is possible to observe the effect of alteringconditions during the experiment. Thus a controlperiod may be followed by a second period during

OXYGEN INLET '\ ! CANNULA

SERUM NEEDLE - --OXYGENOUJTLET

___________SEROSAL FLUID

___ G GUT SAC

POLYTHENE EXTENSION - 1

MMUCOSAL FLUID

FIG. 3. Diagram of the apparatus of Crane and Wilson(1958) as modified by Crampton and Matthews (1964).The wide oxygen outlet prevents blockage by fragments ofdesquamated mucosa, which sometimes occurs in theoriginal apparatus.

which some additional factor expected to influencetransport is introduced (Matthews and Laster 1965b).

In a third technique in vitro, uptake of substancesby the intestinal wall or its substructures is measured.Strips or rings of intestine, sheets of mucosa, andisolated villi have been used (e.g., Agar, Hird, andSidhu, 1954; Crane and Mandelstam, 1960). Suchexperiments give useful information, as in manycases the mechanism of concentration by themucosal cells is an essential part of that for transportacross the intestinal wall. Such techniques have theadvantages that results are more reproducible andthe physical system is simplified; in sac preparationsthe substance transported must diffuse across the

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subepithelial and muscle layers before reaching theserosal fluid.

METHODS in vivo These are of two main types:those in which disappearance of a substance fromthe lumen of the gut is studied, and those involvinganalyses of the blood or lymph.The simplest procedure is to inject a solution of

the substance into tied intestinal loops (e.g., Verzarand McDougall, 1936; Gibson and Wiseman, 1951)and sample the contents at the end of the experiment.As the absorptive capacity of the intestine varieswith site, it is not possible to compare the influenceof different conditions on absorption in differentloops, unless the loops are adjacent and kept veryshort. By cannulation of both ends of a loop ofintestine, the same segment can be used in successiveexperiments under different conditions (Sols andPonz, 1947; Horvath and Wix, 1951), and by con-tinuous perfusion with recirculation from a reservoir,'perfusion in situ' (e.g., Sheff and Smyth, 1955;Fullerton and Parsons, 1951) the concentration ofthe substance being absorbed can be controlled,which is an advantage in experiments on the kineticsof absorption. Whenever such techniques are used,it must be remembered that the information theygive is limited, and that the rate of disappearance ofa substance from the gut lumen does not necessarilyrepresent its rate of entry into the blood or lymph.Substances may disappear from the lumen of the gutnot because of absorption but as the result ofchemical or bacterial action within the lumen. Theymay also be taken up and retained in the wall for anappreciable period, or extensively metabolizedwithin the wall, and some substances disappear fromthe lumen in one form and appear in the blood inanother.

Analyses of the portal and arterial blood duringabsorption also require critical interpretation. Anindwelling portal cannula may be used in theunanaesthetized animal (London, 1935; Dent andSchilling, 1949; Denton, Gershoff and Elvehjem,1953), or the blood may be sampled in an acuteexperiment. In the absence of an estimate of portalblood flow, no quantitative estimate of absorptionrate is obtained. Since alterations in the compositionof the blood during absorption are contributed to bythe tissues, even the qualitative interpretation ofthe data has certain pitfalls (Dent and Schilling,1949; Fisher, 1954). To overcome these difficulties,it is possible to tie off a loop of intestine, cannulatethe vein draining it, and collect all the blood leavingthe loop during the experimental period (Matthewsand Smyth, 1954). The blood is not returned to theanimal, so that the absorbed substance does notappear in the arterial blood supply to the loop. In

this way the total amount of substance absorbedinto the blood can be compared with the amountdisappearing from the intestinal lumen.Many methods which have not contributed

appreciably to the study of the absorption of proteindigestion products have been omitted from thisaccount. More general or more detailed accountshave been published by Crane (1960), Quastel (1961),Smyth (1961b), Wiseman (1961), and Wilson (1962).

THE FORMS IN WHICH PROTEINS ANDTHEIR DIGESTION PRODUCTS ARE ABSORBED

During the digestion of a protein meal, the intestinaltract contains a mixture of amino-acids, peptides,and whole or partially digested proteins, so that it ispossible that any or all of these substances might beabsorbed. The absorption of each group of sub-stances will be dealt with in separate sections,though it may be said at the outset that there islittle doubt that nearly all the ingested proteinenters the blood as amino-acids, regardless of theform in which it leaves the intestinal lumen.

THE ABSORPTION OF AMINO-ACIDS

APPEARANCE IN THE BLOOD There is a large rise inthe individual amino-acids of portal and peripheralblood during absorption of a protein meal (e.g.,Dent and Schilling, 1949; Denton, Gershoff, andElvehjem, 1953; Denton and Elvehjem, 1954; Stein,Bearn, and Moore, 1954; Longenecker and Hause,1959). The amino-acid pattern of this increasebears some relationship to the composition of theprotein fed, but exact correspondence is not to beexpected, as the amino-acid composition of theintestinal contents is not dependent solely on thecomposition of the ingested protein, and is probablysubstantially modified by the hydrolysis of endo-genous proteins derived from enzymes, mucus, anddesquamated epithelial cells (Nasset, 1957). Otherfactors which may modify the amino-acid patternappearing in the blood are differences in the rates ofabsorption of individual amino-acids, and differencesin their rates of uptake by the tissues, which areaffected by cellular requirements and other factors(Longenecker and Hause, 1959).

Fisher (1954) criticized the hypothesis that thebulk of the ingested protein appeared in the blood-stream as amino-acids, pointing out that the rate ofhydrolysis of proteins by known digestive enzymesin vitro was so slow that it seemed unlikely that theycould be completely hydrolyzed to amino-acidsin vivo and absorbed in this form. However, largequantities of amino-acids have been found in the gut

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within a short time of feeding protein (Wright andWynn, 1951), and work in man (Crane and Neu-berger, 1960; Crane, 1961) suggests that protein ishydrolyzed vary rapidly. In spite of this, Fisher'sarguments have received some support from thedemonstration (referred to later) that the finalstages of hydrolysis take place, at least in part, inthe cells of the intestinal mucosa. The idea thatproteins are absorbed into the blood as amino-acidshas also been criticized on the grounds that theconcentration of amino-acids in portal blood duringprotein absorption is too low to account forabsorption entirely in this form. Dent and Schilling(1949) showed that this criticism was invalid for thedog if the large volume of the portal blood flow wasconsidered. In man, if one assumes a portal plasmaflow of 1 litre per minute, and an arterio-portalamino-acid concentration difference of 4 mg.ac-amino N per 100 ml., it can be calculated thatamino-acids equivalent to approximately 15 g. ofprotein per hour could be transported from the gut.It is likely that the portal flow increases duringdigestion, and that the arterio-portal concentrationdifference can exceed 4 mg. when large amounts ofproteins are fed, so that it does not appear that thereis any difficulty in accounting for transport from thegut entirely in the form of amino-acids.

EVIDENCE FOR A SPECIAL ABSORPTIVE MECHANISMMost of the earlier work on the absorption of amino-acids was interpreted as showing that this was due tosimple diffusion (e.g., Chase and Lewis, 1934; An-drews, Johnston, and Andrews, 1936; Kratzer, 1944).Hober and Hober (1937), however, found that theabsorption of amino-acids was much faster than thatof other compounds of similar molecular size, andthat the rate of absorption approached a limitingvalue as the concentration in the intestinal lumenwas increased, and Schofield and Lewis (1947)reported that L-alanine was absorbed more rapidlythan D-alanine. Then Wiseman and others (Elsden,Gibson and Wiseman, 1950; Gibson and Wiseman,1951) showed unequivocally that the L-isomers ofmany amino-acids disappeared from the lumen ofrat small intestine faster than the corresponding D-isomers. A similar phenomenon has also been shownin several species, including the dog, the chicken,and man (Clarke, Gibson, Smyth and Wiseman,1951; Paine, Newman and Taylor, 1959; Kurodaand Gimbel, 1954; Wang and Waisman, 1961).It was also found that the L-isomers were preferen-tially absorbed into the portal blood (Matthews andSmyth, 1952; 1954) which showed that the results ofGibson and Wiseman (1951) could not be accountedfor by bacterial destruction ofL-amino acids within thelumen of the gut, or by their preferential metabolism

within the gut wall. The demonstration of stereo-chemical specificity in absorption strongly suggestedthe existence of a special absorptive mechanism. Evi-dence that the mechanism involved active transportwas presented by Wiseman (1951), who showed thatL-amino acids were transported against a concen-tration gradient by rat small intestine in vitro. TheL-isomers of most of the common monoamino-monocarboxylic amino-acids and of several dibasicamino-acids have now been shown to be activelytransported (e.g., Wiseman, 1953; 1956; Wilsonand Wiseman, 1954; Smyth and Whaler, 1953; Agar,Hird, and Sidhu, 1953; Neil, 1959; Spencer andSamiy, 1960; Wilson, 1962; Wiseman, 1964). Ad-ditional evidence for the existence of special mech-anisms in amino-acid absorption is discussed later.

STRUCTURAL REQUIREMENTS FOR INTESTINAL TRANS-PORT A considerable amount of information hasnow been gained about the molecular structuralfeatures influencing amino-acid transport by thesmall intestine. These features probably reflect someofthe chemical properties of the transport mechanismor mechanisms. So far attention has been paidmainly to the requirements for active transport.The structure of a typical amino-acid is:

H

R-C-COOH

NH2

A carboxyl group is attached to a carbon atom,designated the ac-carbon. To this a-carbon atomare attached a hydrogen atom, an amino group, anda side chain, 'R'. This side-chain takes variousforms, which distinguish the individual amino-acids.It may be a simple aliphatic chain, straight orbranched (e.g., alanine, valine, leucine), it maycontain sulphur atoms (e.g.., methionine, cysteine),or it may include an aromatic or heterocyclicring (e.g., phenylalanine, tyrosine, histidine, trypto-phan). It may include a second carboxyl group,forming a dicarboxylic amino-acid (e.g., glutamicacid, aspartic acid) or a second amino group,forming a dibasic (diamino) amino-acid (e.g.,lysine, arginine, ornithine). All amino-acids whichhave four different groups around the oc-carbon canoccur in L- or D- forms according to the dispositionof the groups, these stereoisomers being mirrorimages of each other, and similar in most physicaland chemical properties. The L-isomers are some-times known as the 'natural' isomers since they arethe only forms commonly met with in foodstuffsand in the tissues of higher animals. In amino-acidsin which there are only three different groups

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attached to the ac-carbon, these optical isomerscannot occur; thus glycine, in which R is representedby a second hydrogen atom, has only one form.The following structural features are important in

active transport (e.g., Wiseman, 1955; 1956;Wilson, Lin, Landau, and Jorgensen, 1960; Christen-sen, 1960; Spencer, Bow, and Markulis, 1962; Lin,Hagihira, and Wilson, 1962; Wiseman, 1964).

The amino group This is essential for activetransport. Thus, acetic acid, which may be regardedas glycine in which the amino group has beenreplaced by a hydrogen atom, is not activelytransported. Replacement of one of the hydrogenatoms of the ac-amino group, as in proline andN-methylglycine, does not necessarily prevent activetransport, though it may do so if the substitutinggroup is bulky, as in N-phenylglycine. If bothhydrogen atoms are substituted, as in N-dimethyl-glycine, active transport is abolished. It also appears

to be essential for the amino-group to be attachedto the ac-carbon atom; if not, as in /-alanine, activetransport will not take place.

The carboxyl group The carboxyl group alsoappears to be essential, and it must be free; thus themethyl ester of L-histidine is not actively transported.

The hydrogen ofthe ac-carbon This is not essential,since the amino-acids ac-aminoisobutyric acid,isovaline, and cycloleucine (which are apparentlynot metabolized and in which the ac-hydrogen isreplaced by a methyl or other group, are activelytransported, but there is evidence that the substitutionreduces affinity for the transport mechanism(Christensen, 1962).

Stereochemical configuration at the ac-carbonUntil recently, it was thought the L-configurationwas an absolute requirement for active transport,but the finding (Jervis and Smyth, 1960) thatD-methionine is transported against a concentrationgradient suggests that this is not a universal rule.Evidence from experiments on competition fortransport and on the kinetics of the transportprocess also indicates that the absorption of D-amino-acids, though it may be passive in manycases, is unlikely to be by simple diffiusion.

The side-chain The nature of the side-chain mayalso determine whether or not an amino-acid isactively transported. In the case of the mono-

aminomonocarboxylic amino-acids, the length ofthe side-chain appears to have an important influenceon affinity for the transport mechanism. For some

time it was thought that only the monoaminomono-carboxylic amino-acids were actively transported,but it has since been shown that the diamino acids,lysine, ornithine, and arginine, are also transportedactively. There is no evidence for active transportof the dicarboxylic acids, glutamic and aspartic,

though possibly such evidence is masked by extensivemetabolism of these compounds in the intestinalwall.

COMPETITION FOR INTESTINAL TRANSPORT If twocompounds compete for transport, this suggeststhat they share at least one stage in the transportprocess. The first systematic study of competitionfor absorption between amino-acids was that ofWiseman (1955). He found that diamino anddicarboxylic amino-acids had no effect on thetransport of several monoaminomonocarboxylicamino-acids. Among the monoaminomonocarboxylicamino-acids, those transported at a slow rate wereparticularly effective in inhibiting the transport ofthose transported at a faster rate. Thus methionine,which is relatively slowly transported at a con-centration of 20 mM, almost completely preventedtransport of proline, glycine, and histidine, whereasglycine, which is relatively rapidly transported atthis concentration, had little or no inhibiting effecton transport of the more slowly transportedhistidine and methionine. As a result, he suggestedthat the monoaminomonocarboxylic amino-acidsshared a common transport mechanism, in whichthe slowly transported amino-acids had a highaffinity for some part of the mechanism and formeda comparatively stable union with some part of it,thus accounting for their slow transport and highinhibiting ability.

These and other observations now enable furthergeneralizations to be made. It is probable that theL-isomers of most, if not all, monoaminomono-carboxylic amino-acids compete with each other forintestinal transport (e.g., Pinsky and Geiger, 1952;Fridhandler and Quastel, 1955; Agar, Hird andSidhu, 1956; Finch and Hird, 1960b,. Hagihira,Ogata, Takedatsu, and Suda, 1960; Spencer andSamiy, 1961), and that those with long side-chainsare the most powerful inhibitors of the transport ofother amino-acids within this group. The longer theside-chain, the more powerful the inhibiting effect,i.e., the greater the apparent affinity for the commontransport mechanism (Finch and Hird, 1960b;Nathans, Tapley, and Ross, 1960; Matthews andLaster, 1965b). It also appears that there are severalintestinal transport mechanisms for different groupsof amino-acids, there being competition among themembers of each group, but not between the mem-bers of different groups:

1 The mechanism shared by the majority ofmonoaminomonocarboxylic amino-acids; 2, amechanism for the transport of the three basicamino-acids, ornithine, lysine, arginine, and cystine(Hagihira, Lin, Samiy, and Wilson, 1961); 3, amechanism for the transport of N-substituted amino-

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acids (betaine, dimethylglycine, and sarcosine).Two other N-substituted amino-acids, proline andhydroxyproline, appear to be transported partly bythis system and partly by that for the monoamino-monocarboxylic acids (Wilson, 1962; Hagihira,Wilson and Lin, 1962). 4 Possibly a specialmechanism for the transport of glycine: this issuggested by the work of Christensen (1962) andAkedo and Christensen (1962), and Newey andSmyth (1963) have shown that glycine is probablytransported partly by a special mechanism ofits own and partly by that for the other mono-aminomonocarboxylic amino-acids. All these experi-ments, however, were carried out with rat gut. In thehamster, we could find no evidence that glycine wasnot transported entirely by mechanism (1), so thatthere may be species differences in the transport ofthis amino-acid (Matthews and Laster, 1965b).The results of animal experiments and of investi-

gation of amino-acidurias in man suggest that thereare similar groups of transport mechanisms in thekidney, and that certain defects in the renal mechan-isms, as in Hartnup disease and cystinuria, areaccompanied by corresponding defects in theintestinal mechanisms (see Milne, 1964).

Competition for intestinal transport is notconfined to L-amino acids, but apparently alsooccurs between some D-amino acids (Agar et al.,1956). Competition has also been observed betweenL-amino acids and D-amino acids (Jervis andSmyth, 1959b; Paine et al., 1959). This raises thepossibility that there is at least one stage in transportwhich is common to both L- and D- isomers.

THE KINETICS OF INTESTINAL TRANSPORT Additionalinformation about the mechanisms of intestinaltransport can be gained by studying their kinetics.

It has already been pointed out that if a substanceis absorbed by simple diffusion, its rate of absorptionshould be approximately proportional to its con-centration within the intestinal lumen. If, on theother hand, a special mechanism (active transportor facilitated diffusion) is involved, the absorptionrate will not keep pace with increasing concentration,but will approach a limiting value as the absorptivemechanism becomes saturated. Hober and H6ber(1937), Lathe (1943), and Hetenyi and Winter(1952) observed that the absorption of amino-acidstended to approach a limiting rate, and in recentyears the kinetics of amino-acid absorption havebeen investigated in more detail, by techniquesboth in vivo and in vitro.

If the rates of transport of amino-acids areplotted against their concentrations in the intestinallumen or on the mucosal side of a sac preparation,curves of the type shown in Fig. 4 are usuallyobtained. Transport rate increases with increasingconcentration until a maximal rate is attained, thisrate varying according to the individual amino-acid.If the reciprocal of the transport rate at eachconcentration is plotted against the reciprocal of theconcentration (excluding the highest concentrations),the result is a straight line (Fig. 5). Figure 5 isanalogous to the Lineweaver-Burk plot of enzymekinetics, and this linear relationship has been takento mean that the transport process conforms to theMichaelis-Menten kinetics of enzyme reactions.Kinetics of this type, which have now been shownfor a number of L-amino acids and two D-aminoacids (e.g., Jervis and Smyth, 1959a; Finch and Hird,1960b; Nathans et al., 1960; Spencer and Samiy,1961; Lin et al., 1962; Matthews and Laster, 1965a),are to be expected if the rate-limiting step in transportis combination with or adsorption to some com-

FIG. 4. The relationship between initialamino-acid concentration in the mucosalfluid and amino-acid transport into theserosal fluid, using sacs ofhamster smallintestine. A-glycine, o-L-alanine,*-L-valine, E-L-leucine, EII-amino-isobutyric acid. The tendency for the ratesto decline at very high concentrations maybe characteristic of the experimental

~~-a system. (Matthews and Laster, 1965a;reproduced by permission of the Editor ofthe American Journal of Physiology).

INITIAL AMINO ACID CONCENTRATION CmM)

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0o005

0-004-

0-003-

1/vo-oo2 -

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caiofThin

posizmetrawi1dasys19'reswech;to

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TABLE ICHEMICAL STRUCTURE AND TRANSPORT CONSTANTS FOR

A SERIES OF NEUTRAL AMINO-ACIDS1

Amino-acid Side-chain Kt (mM.) Vmax (AM/S-dry wt./hr.)

Glycine H- 43L-alanine CH,- 7-5L-valine (CH,),CH- 19L-leucine (CH,),CH.CH,- 1-5

'Modified from Matthews and Laster (1965a)

2,6001,700960490

chains are powerful competitive inhibitors of

/-001- transport, and thus appear on these grounds to have0001§ - D/ a high affinity for the transport mechanism. Further,

using the Kt values obtained from kinetic experi-ments, and making certain assumptions, it ispossible to calculate the expected degree of inhibition

62 0 0'-2 0-4 0-6 0-8 of transport of one amino-acid when it is transported

lC in the presence of another (Finch and Hird, 1960b;. 5. Lineweaver-Burk plot for L-alanine r

Matthews and Laster. 1965b). In a limited series ofrpc monoaminomonocarboxylic amino-acids, the pre-

of amino-acid concentration (mM) = reciprocal dicted values agree reasonably well with thosecorresponding transport rate (,umoles/g. dry wt./hr.). actually observed in competition experimentse data from which the plot was constructed are included (Table II). Lin et al. (1962) have proposed a hypo-Figure 4. thesis to explain the relationship between chemical

ynent of the cell membrane, but it must be empha- structure and transport characteristics. They suggeste that the more lipophilic the side-chain, the moreted that such kinetics do not show that a carrier reat t more in the sid-chain,membrezchanism or even an enzyme system is involved in eadlyitdissolves in the lipid-rich cell membraneinsport. The findings must in fact be interpreted permitting the carboxyl group, amino group andth caution, because any such treatment of the a-hydrogen to reach the carrier or carrier site, whichLta involves gross oversimplification Of a complex hsatcmnsfrteegop.Ti olstem (Wilbrandt and Rosenberg, 1961; Wolff+ explain the association between length of side-chain64; Matthews and Laster, 1965a). In spite of these and low Kt values. They also suggest that the reason64; Matosit is possible to derive from the Line- why highly lipophilic side-chains are associated with*ervations, pos to dere om traneo low maximal transport rates is that they do not

aver.Burk plotstwo measures of transpot. readily dissociate from the cell membrane, so thataracteristics which show a definite relationship transport is impeded. The results so far obtained arechemical structure (Finch and Hird, 1960b; cnitn ihti yohssatthews, and Laster 1965a). The reciprocal of the consistent with this hypothesis.attancews andfrom stheorin1 .The rec ca l ofthena Before leaving the subject of kinetics, it shouldtance Y from the origin (Fig. 5) may be taken as be noted that it is possible for an amino-acid A to belicating the apparent limiting velocity of transport transported faster than an amino-acid B at one

111e1-,ctpmicl;iU V )Y maxJnthtu"' nr nf tht-

distance X as indicating the concentration at whichthe transport rate is half-maximal (Kt). Low Ktvalues indicate that it is relatively easy to saturatethe transport mechanism with the substance trans-ported, in other words, low Kt values suggest highaffinities for the transport mechanism, and high Ktvalues suggest low affinities.

Table I shows a series of monoaminomono-carboxylic amino-acids, and the correspondingVmax and Kt values obtained from sac experiments.As the length ofthe side-chain increases, Kt decreases,i.e., the apparent affinity for the transport mechanismbecomes greater, and Vmax also decreases. Theseresults agree well with those of competition experi-ments, which show that amino-acids with long side-

TABLE IICOMPETITIVE INHIBITION OF AMINO-ACID TRANSPORTINTO THE SEROSAL FLUID OF EVERTED SACS OF

HAMSTER SMALL INTESTINE'Inhibited No. of Inhibitor Concen- Percentage InhibitionAmino-acid Experi- Amino- tration of of Transport(concen- ments acid Inhibitortration Amino- Observed Predicted20 mM.) acid (mM)

Glycine 2 L-alanine 52 L-alanine 203 L-leucine 2

L-alanine 2 Glycine 403 L-leucine 2

L-leucine 2 Glycine 402 L-alanine 40

'Modified from Matthews and Laster (1965b)

31 3186 6462 4721 2030 260 6

27 28

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concentration, but more slowly at another (Fig. 4),and according to the concentration used, a series ofamino-acids may show several different orders oftransport rate. This may explain why in earlierinvestigations where amino-acid absorption wasstudied at one concentration only (e.g., Kratzer,1944), no clear relationship emerged betweentransport rate and chemical structure. There is,however, a general tendency for amino-acids withlow Kt values to be transported relatively fast atlow concentrations, and relatively slowly at highconcentrations. This probably explains why Wise-man (1955), working at the comparatively highconcentration of 20 mM, found a correlationbetween slow transport and powerful competitiveability. It is now clear that to say that a particularamino-acid is transported relatively 'fast' or 'slowly'is meaningless unless the concentration isspecified.

RELATIONSHIP OF INTESTINAL TRANSPORT TO META-

BOLIC ENERGY SUPPLY There is little doubt thatthe energy for the active transport of amino-acidsby the intestine of the adult animal is derived fromoxidative metabolism. Transport is inhibited byanoxia and various metabolic inhibitors, includingcyanide and 2: 4 dinitrophenol (e.g., Agar et al.,1953; Wilson and Wiseman, 1954; Jervis andSmyth, 1960). There remains, however, a possibilitythat the mechanisms affected are not in fact directlyconcerned in supplying energy for transport butserve primarily to maintain the integrity of the cellmembrane, without which the transport processcannot operate (Finch and Hird, 1960a; Newey andSmyth, 1962). The energy supply is probably depend-ent on adenosine triphosphate (A.T.P.). It has beenclaimed (Feher, Kertai, and Gati, 1956; Kertai,Feher, and Gati, 1956) that if resynthesis of A.T.P.is blocked by iodoacetic acid, the A.T.P. content ofthe intestinal wall falls during the absorption ofglycine, and that when the wall is depleted of A.T.P.,the rate of glycine absorption diminishes. However,these authors found that the A.T.P. content alsofalls during the absorption of urea, which is almostcertainly not an active process. This raises somedoubt about the significance of their findings.According to Shishova (1956; 1959), the presence ofA.T.P., magnesium, and phosphate in the intestinallumen accelerates the absorption of amino-acids,though Christensen (1960) comments that anenergizing action of A.T.P. added to the outside of atissue is very unusual. Unlike the adult gut, smallintestine of foetal and newborn rabbits can transportamino-acids actively under anoxic conditions, theenergy apparently being supplied by anaerobicglycolysis (Wilson and Lin, 1960).

MISCELLANEOUS FACTORS INFLUENCING AMINO-ACIDABSORPTION These must be considered separately.

Pyridoxal and its derivatives; Other vitaminsOwing to the suggestion that a derivative of pyri-doxal might function as a carrier in the mechanismfor amino-acid concentration by tissue cells (Christen-sen, Riggs, and Coyne, 1954), several studies havebeen made on the effect of this compound and otherforms of vitamin B6 on amino-acid absorption.Though the addition of pyridoxal has no effect onthe transport of amino-acids by hamster smallintestine in vitro (Wiseman, 1957), amino-acidabsorption is depressed by pyridoxal deficiency andby the antimetabolite desoxypyridoxine (Fridhandlerand Quastel, 1955; Jacobs and Hillman, 1958).Inhibition of transport produced by 2: 4 dinitro-phenol is said to be reversible by pyridoxal phosphatebut not by pyridoxal, and it has been suggested thatthe effect of D.N.P. is to interfere with the phosphory-lation of pyridoxal, thus cutting off the supply ofpyridoxal phosphate, which is supposed to be activein the absorptive mechanism (e.g., Jacobs, Coen,and Hillman, 1960). While there is little doubt thatpyridoxal and related compounds have someinfluence on the absorption of amino-acids, theirrole in a carrier mechanism is more than doubtfuland their mode of action still not clear. Perhapspyridoxal modifies the cell membrane so as to slowthe efflux of amino-acids (Christensen, 1962). Themembers of the B6 group are the only vitamins whichhave definitely been shown to affect amino-acidabsorption, though Erf and Rhoads (1940) notedthat the glycine tolerance curve was depressed inpernicious anaemia but not in other severe anaemias.Any effect of B12 deficiency may well be non-specific (Reynolds, Hallpike, Phillips and Matthews,1965). The absorption of xylose may also be defec-tive in pernicious anaemia (e.g., Butterworth, Perez-Santiago, de Jesus, and Santini, 1959).

Electrolyte concentration The active transport ofamino-acids by the small intestine is sodium-dependent, and, in the absence of this ion, is greatlyreduced or abolished. This is not a specific effect,as the transport of water, glucose, and inorganicphosphate is also inhibited. Low concentrations ofpotassium have no effect (at least on monoiodo-tyrosine transport) but high concentrations areinhibitory, and the inhibition appears to be competi-tive. Alterations in calcium concentration have noeffect (Nathans et al., 1960; Csaky, 1961; Harrisonand Harrison, 1963).

Water absorption Using in vitro preparationsthere is a correlation between water and amino-acidtransport (Matthews and Laster, 1965a) as there isbetween water and glucose transport (Fullerton andParsons, 1956; Smyth and Taylor, 1957). Possibly

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the absorption of amino-acids is assisted by 'solventdrag', that is, the entrainment of solute moleculesin a water stream. There are, however, several otherpossible explanations for this association, and thework of Newey and Smyth (1962) suggests that sol-vent drag is not an important factor in the absorptionof amino-acids.pH The optimal pH for transport of monoiodo-

tyrosine by rat small intestine in vitro is about 6(Nathans et al., 1960).

Bile Absence of bile from the intestine is said todecrease nitrogen absorption (see Wiseman, 1964).Dawson and Matthews (1961) found that theaddition of conjugated bile salts to sac preparationsincreased the permeability of the cells of theintestinal wall to amino-acids, allowing largerquantities to pass from the interior of the cells intothe surrounding medium, but there was no appreci-able effect on amino-acid transport from mucosalto serosal fluids. On the other hand, Parkinson andOlson (1963) found that transport was reduced, andsuggested that bile salts might 'regulate' the absorp-tion of various nutrients. Unconjugated bile saltsabolish the active transport of amino-acids andglucose, and damage the intestinal mucosa (Dawsonand Isselbacher, 1960).

Diet Kershaw, Neame, and Wiseman (1960)found that if rats were given a restricted diet for afew days there was a striking increase in the rates ofabsorption of L-histidine and of glucose, both invivo and in vitro, in spite of the fact that the smallintestine became shorter, narrower, and thinner-walled. The increased absorptive ability disappearedafter several days on a normal diet. A similarincrease in absorption rate was found in sarcoma-bearing rats (Wiseman, Neame, and Ghadially,1959). The mechanisms of these adaptations areunknown. Lin and Wilson (1960) gave rats additionaltyrosine for a month, but found no change in itsabsorption rate.

Monosaccharide absorption There are conflictingreports about the effects of monosaccharides onamino-acid absorption. Cori (1926) reported thathigh concentrations of glucose in the intestineinhibited the absorption of amino-acids, an effectwhich Fridhandler and Quastel (1955) felt wasprobably osmotic. Agar et al. (1953) stated thatglucose and amino-acids did not compete forabsorption. More recently Newey and Smyth (1964)reported that in sac preparations glycine transportis stimulated by glucose but inhibited by galactose,and consider that this may be due to alterations inthe amounts of A.T.P. available for supplying energyfor amino-acid transport.

Drugs Little is known about the effects of drugson amino-acid absorption. Phlorrhizin, which

inhibits the absorption of glucose, probably byinterfering with its entry into the mucosal cells, hasno effect on the absorption of amino-acids (Frid-handler and Quastel, 1955; Parsons, Smyth, andTaylor, 1958). Chlortetracycline in the intestinallumen inhibits the absorption of L-histidine (Agarand Parker, 1958); the authors suggest that this isdue to interference with energy supply owing todepression of mitochondrial phosphorylation.Draper (1958) claimed that oral penicillin increasedthe absorption of lysine. The effect of anaesthesiaon amino-acid absorption does not seem to havebeen studied.

Species differences Most of the recent work onamino-acid absorption has been carried out withintestine from the rat or golden hamster, but observ-ations have also been made in the frog, chick,rabbit, cat, dog, guinea-pig, and in man. No majordifferences in the absorptive mechanisms have yetbeen observed, though the possibility of minordifferences has already been referred to. Lin andWilson (1960) compared the absorption rates ofL-tyrosine in the chick, hamster, rabbit, and rat:the rates decreased in that order.

Other possible influences on amino-acid absorptionMany factors that may influence the absorption ofamino-acids have not yet been investigated. Thereis very little information about the effects ofalterations in intestinal motility or in blood flow,though Cummins and Almy (1953) reported thatdrug-induced hypermotility accelerated methionineabsorption. Nothing is known about the effects ofhormones, nor about the effects of age or sex.(Sex differences are believed to exist in the rate ofabsorption of glucose.)

METABOLIC CHANGES DURING ABSORPTION Numerousmetabolic changes have been shown to occur duringthe intestinal transport of amino-acids, but there isno positive evidence that they are related to thetransport mechanisms, and they are, with a fewexceptions, quantitatively unimportant (Wiseman,1953; Whaler, 1955; Finch and Hird, 1960a;Wiseman, 1964). The possible metabolic pathwaysavailable to amino-acids in the intestinal wall aresummarized by Spencer and Knox (1960).

Substantial amounts of L-glutamic and L-aspartic acids are transaminated during absorption(Matthews and Wiseman, 1953; Neame and Wise-man, 1957; 1958) which results in the appearanceof alanine in the portal blood, in addition to theamino-acid present in the intestinal lumen. When thelumen concentration of glutamic acid is low, it isall transaminated, and only alanine appears in theblood. Many metabolic pathways are apparentlyavailable to glutamine and glutamic acid in the

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intestinal wall (Spencer and Knox, 1960; Spencerand Zamcheck, 1961) and it is potentially able tointerconvert these compounds, but there is noevidence that such interconversion occurs duringabsorption, at least in the cat and guinea-pig(Bessman, Magnes, Schwerin, and Waelsch, 1948;Fridhandler and Quastel, 1955). In contrast toglutamine, asparagine is almost completely hydro-lysed to aspartic acid and ammonia during transportby guinea-pig intestine (Fridhandler and Quastel,1955). Sugawa, Akedo, and Suda (1960) report thatmethionine sulphoxide is formed during the absorp-tion of methionine by the rat, and partly reducedagain to methionine in the portal blood.The small intestine, which is constantly producing

fresh epithelial cells, enzymes, and mucus, is anextremely active site of protein synthesis (Block,1946; Friedberg, Tarver, and Greenberg, 1948;Busch, Davis, Honig, Anderson, Nair, and Nyhan,1959) and it is quite possible that small amountsof amino-acids are diverted to protein synthesisduring the process of absorption. Labelled aminoacids given orally appear very rapidly in the proteincoating of chylomicrons, which suggests that thismay be synthesized in the intestinal wall (Bragden,1958).

THE SITE AND ROUTE OF ABSORPTION Many referencesto early work on the site of absorption of proteindigestion products are given by Verzar andMcDougall (1936). The more recent observations,which have mostly been made on sacs of smallintestine from the rat or hamster, show that the siteof maximum ability to transport monoaminomono-carboxylic amino-acids against a concentrationgradient is in the ileum, in the mid to lower part ofthe small intestine (e.g., Wilson and Wiseman, 1954;Lin and Wilson, 1960; Spencer and Samiy, 1960;1961; Nathans et al., 1960; Christensen, 1962;Matthews and Laster, 1965a). These findings are incontrast to those for monosaccharides, which arebest transported in the duodenum (see Crane, 1960).L-cystine and L-cysteine may be unusual in beingbest transported in the duodenum and jejunum(Neil, 1959). There is probably no active transportmechanism for amino-acids in the colon (Corderoand Wilson, 1961).

It is not yet clear what determines the site ofmaximal absorptive capacity for a particularsubstance. The findings quoted cannot be explainedmerely as a function of mucosal surface area, or ofthe rate of aerobic metabolism, both of which aregreatest in the upper small intestine. It should beemphasized here that the site of maximum absorptiveability as determined in vitro is not necessarilyidentical with the site in which most absorption takes

place in vivo (Crampton and Matthews, 1964). It is atleast possible that with a substance which is veryreadily absorbed, absorption may be virtually com-plete before the lower part of the small intestine isreached.Having left the cells of the mucosal epithelium,

amino-acids pass across the subepithelial space intothe capillaries of the villi, presumably by diffusion.While there is no doubt that the main route ofabsorption is into the blood, absorption into thelymph is not necessarily entirely negligible (Hendrixand Sweet, 1917; Bolton and Wright, 1937). It isassumed that absorption into the blood predominatesbecause the rate of blood flow is far greater thanthat of the lymph (see Wiseman, 1964) and becauseof the peripheral position of the capillaries of thevilli as opposed to the central position of the lacteals.

THE ABSORPTION OF PEPTIDES

There has never been any satisfactory evidence thatpeptides are absorbed into the blood in appreciablequantities. The older claims that large amounts ofpeptides appear in the blood after a protein meal(e.g., London and Kotschneff, 1934) may be dis-counted on the grounds of inadequate analyticaltechnique (Christensen and Lynch, 1946; Christen-sen, Decker, Lynch, MacKenzie, and Powers, 1947),and more recent investigations have shown thatthere is no significant increase in blood peptides oramino-acid conjugates after protein feeding (Dentand Schilling, 1949; Parshin and Rubel 1951;Christensen et al., 1947; Stein and Moore, 1954).However, the advent of in vitro methods has thrownnew light on the question of the absorption ofpeptides and revealed some unexpected facts.When the transport of di- and tripeptides was

investigated using isolated small intestine, it wasfound that (with the exception of small quantities ofglycylglycine) they were unable to cross the intestinalwall intact; they were hydrolysed by the preparation,and only their constituent amino-acids appeared onthe serosal side of the gut (Agar et al., 1954;Johnston and Wiggans, 1958; Wiggans and John-ston, 1959). Newey and Smyth (1959; 1960) investi-gated the question of whether the failure of smallpeptides to cross the intestinal wall was due tohydrolysis in the fluids surrounding the gut, or tohydrolysis within the intestinal wall. Using bothin vitro and in vivo methods, they showed thatextracellular peptidases were capable of hydrolysingonly a small fraction of the peptides disappearingfrom the mucosal side of the intestine; this meantthat the peptides must be hydrolysed, either at thesurface of the mucosal cells, or within them. Furtherexperimental evidence indicated that most of the

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hydrolysis was intracellular. Subsequent work(Newey and Smyth, 1962), carried out mainly withglycylglycine, suggests that dipeptides enter thecells of the intestinal mucosa partly by diffusion,but not entirely so, since the rate of entry of glycyl-glycine is approximately the same as that of glycine,and entry is reduced by anoxia. They are hydrolysedwithin the cells, and the amino-acids formed attachthemselves to a transport mechanism within thecells (though there is some diffusion back into theintestinal lumen). This transport mechanism, whichNewey and Smyth term the 'exit' mechanism, may beshared by free amino-acids which have entered fromthe lumen of the intestine.

This work shows that in addition to the absorptionof free amino-acids, there is probably a second modeof absorption of protein digestion products, in whichsmall peptides enter the mucosal cells, are hydrolysedwithin them, and pass into the blood stream asamino-acids. The proportions of ingested proteinwhich leave the intestinal lumen in the form of freeamino-acids and in the form of peptides are not yetknown. It is interesting that these recent investiga-tions show that there was some truth in the old view(Starling, 1906) that the final hydrolysis of food-stuffs might occur during the actual process ofabsorption.

THE ABSORPTION OF WHOLE PROTEINS

In the first few days or weeks of life, animals ofseveral species can absorb a variety of proteinsapparently unchanged, and substantial amounts ofcolostrum proteins, including maternal antibodies,are absorbed in this way (e.g., Howe, 1921; Smith,1948; McCance, Hutchinson, Dean, and Jones, 1949;Hill and Hardy, 1956; Clark, 1959; McCance andWiddowson, 1959). The process is selective, forforeign proteins are absorbed less well than maternalantibodies, and interfere with their absorption(Brambell, Halliday, and Morris, 1958). Duringthe absorption of whole protein, it appears invacuoles within the columnar cells of the intestinalmucosa. These are formed by invagination of theapical cell membrane, i.e., absorption is by pino-cytosis. The ingestion of materials in this way ischaracteristic of certain cells in primitive organisms,and its persistence in the gut of the newborn animalprobably serves a nutritive function while thedigestive mechanisms are still relatively undeveloped(Hill, 1956). In experimental animals, the ability toabsorb whole protein is lost after the first two tothree weeks, and this can be hastened by givingcortisone (Halliday, 1958; Clark, 1959). In rats andmice, the site of absorption of whole proteins is the

jejunum and ileum, but not the duodenum (Clark,1959); the route of absorption is via the lymphaticsand not into the blood (Comline, Roberts, andTitchen, 1951). Comparatively little is known aboutthe absorption of whole proteins in the humanneonate, but there is no evidence for substantialabsorption of maternal antibodies (Kekwick, 1959).Babies can be shown to absorb small amounts ofintact cow's milk, egg, and other foreign proteins byimmunological techniques (e.g., Wilson and Walzer,1935; Gruskay and Cooke, 1955) but this does notindicate that whole proteins are absorbed innutritionally significant amounts.

It is very unlikely that adult animals can absorbintact foreign proteins except in minute quantities.Rats can be sensitized to foreign proteins givenorally (Ratner and Gruehl, 1934), and after givingegg albumin to dogs, it is detectable by a precipitinreaction in the lymph, though not in the portalblood (Alexander, Shirley, and Allen, 1936). Allergicreactions to a variety of food proteins are notuncommon in man, and are evidence of absorptionof protein either unchanged or only slightly modified;such absorption can also be shown immunologically(e.g., Sussman, Davidson, and Walzer, 1928). In allthese examples, absorption is on a very small scale.Under certain conditions, insulin is absorbed fromintestinal loops in dogs (Murlin, Tomboulian, andPierce, 1937); it is also absorbed by rats if proteo-lysis is artificially inhibited, and can cross the wallsof intestinal sacs (Danforth and Moore, 1959), butin these experiments the normal digestive processesare prevented.The literature contains several reports suggesting

that plasma proteins from an animal itself or fromanother of the same species may be absorbed inlarge quantities either intact or at any rate withoutextensive hydrolysis (Voit and Bauer, 1869; Heiden-hain, 1894; Visscher, Roepke, and Lifson, 1945;Dent and Schilling, 1949). Dent and Schilling fedhomologous plasma to dogs, and it appeared to beabsorbed without significant alteration in the amino-acids or small peptides of the portal blood. Thiscould mean that either it was absorbed intact, or inthe form of peptides too large to be detected by thepaper chromatographic methods used. Subsequentwork, however, suggests that homologous plasmaproteins are completely hydrolysed during absorp-tion (Abdou and Tarver, 1951; Yuile, O'Dea,Lucas, and Whipple, 1952).

HYPOTHESES OF THE MECHANISMS OFABSORPTION OF AMINO-ACIDS AND PEPTIDES

It appears from work with small intestine in vitrothat concentrative uptake of amino-acids by the

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Absorption ofprotein digestion products: A review 423

cells of the intestinal mucosa is an essential part ofthe process of active transport across the intestinalwall (Agar et al., 1956; Finch and Hird, 1960a;Newey and Smyth, 1962). For this reason, andbecause there are many resemblances betweenintestinal transport of amino-acids and theirtransport into tissue cells, Christensen (1960; 1962)has proposed that the intestinal transport mechanismis essentially a modification of the ordinary cellconcentration mechanism. If transport into themucosal cells were more vigorous on the mucosalside than on the other, transport across the cellswould take place. Christensen and Oxender (1960)have shown experimentally that this is possible, byforming Ehrlich ascites carcinoma cells into a thinmembrane on a fine filter. If uptake is stimulated onone side of this membrane (by adding pyridoxal orestablishing a potassium gradient) amino-acids aretransported against a concentration gradient fromone side to the other. In the intestine, it is possiblethat more vigorous transport on the luminal borderof the mucosal cells might be produced by anextra number of transport sites on this side, perhapssituated in the brush border.The work of Newey and Smyth suggests that

transport across the mucosal epithelium is morecomplicated than this (Smyth, 1961a; Newey andSmyth, 1962). They propose that there are twostages in the transport of glycine: (1) an entrymechanism, where competition with methionineoccurs, and (2) an exit mechanism, which is relativelyuninfluenced by methionine competition, but doesrequire aerobic energy. This exit mechanism isprobably shared by amino-acids which have beenliberated within the cells by hydrolysis of peptides.The nature of the mechanisms responsible for

amino-acid transport across cell membranes isunknown. Certain chemical reactions which mightpossibly be concerned in amino-acid transport intotissue cells have been ruled out by the observationthat there is no exchange of carboxyl oxygen duringthe transport process (Christensen, Parker, andRiggs, 1958). The glucuronide formation that hasbeen reported during the intestinal transport ofmonoiodotyrosine (Tapley, Herz, Ross, Deuel, andLeventer, 1960) and the formation of methioninesulphoxide during the absorption of methionine(Sugawa et al., 1960) may well be incidental ratherthan reactions concerned in the transport mechanism.The hypothesis of the 'shuttling carrier' has alreadybeen touched on. For a time it appeared that thecarrier involved in amino-acid transport might bepyridoxal or some related compound, but this nowseems unlikely. Christensen (1962) points out that inspite of the usefulness of the carrier concept, noexample of a carrier has yet been identified either in

the intestine or elsewhere; he discusses other possibleexplanations of carrier-like behaviour in membranetransport. The terms 'carrier' and 'carrier site' nowtend to be used merely to refer to hypotheticalreactive sites concerned with transport, withoutcommitting the user to the shuttling carrier concept.

Certain characteristics of the 'carrier sites'involved in intestinal transport of amino-acids areindicated by the structural requirements for trans-port, and these have been outlined. The fact thatdifferent groups of amino-acids appear to be trans-ported by separate mechanisms suggest that thereare several categories of transport site with differentchemical specificities.The details of how the metabolic energy supply is

linked to the transport mechanism or mechanismsare unknown. Christensen et al. (1954) suggestedthat in tissue cells, amino-acid transport might belinked with, and perhaps driven by, potassiumtransport. A raised potassium concentration acts likea competitive inhibitor of amino-acid transport(Nathans et al., 1960) which suggests that the twotransports are also associated in some way in theintestine. The abolition of the transport of bothglucose and amino-acids by low sodium concentra-tions may be the result of interference with acommon energy supply to the two mechanisms.Crane (1960) proposed that the active transport ofsugars by the intestine might be driven by theextrusion of sodium from the base of the microvilli,leading to an inflow of water carrying solutes with it,the entry of solutes being controlled by the chemicalspecificities of special entry sites. Possibly a similarmechanism might be invoked to account for theactive transport of amino-acids.Though there have been many advances in the

last 15 years, it is not yet possible to form a completepicture of the intestinal absorption of proteinbreakdown products, and one of the centralproblems, the nature of the transport mechanismsfor amino-acids, is still unsolved. Much remainsto be learnt, and recent physiological findings haveonly just begun to be applied in clinical investigation.The field continues to offer great opportunities.

REFERENCES

Abderhalden, E (1912) Synthese der Zellbausteine. Springer,Berlin.Korbsy. K V., and London, E. S. (1907) Weitere Studien uberdie normale Verdau65ng der Eiweisskbrper im Magendarm-kanal des Hundes. Hoppe-Sevlers Z. physiol. Chem., 53,148-163.

Abdou, 1. A., and Tarver, H. 11951). Plasma protein. 1. Loss fromcirculation and catabolism to carbon dixoide. J. biol. Chem.,190, 769-780

Abel, J. J., Rowntree L. G., and Turner B. B. (1914). On the removalof diffusible substances from the circulating blood of livinganimals by dialysis. J. Pharmacol. exp. Ther., 5, 275-316.


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424 D. M. Matthews and Leonard Laster

Agar, W. T., Hird, F. J. R., and Sidhu, G. S. (1953). The activeabsorption of amino-acids by the intestine. J. Physiol. (Lond.),121, 255-263.

-,-, - (1954). The uptake of amino-acids by the intestine.Biochim. biophys. Acta (Amst.), 14, 80-84.

-, -, - (1956). The absorption, transfer and uptake ofamino-acids by intestinal tissue. Ibid., 22, 21-30.

, and Parker, M. A. (1958). The inhibition of the intestinalabsorption of an amino-acid by chlortetracycline. Ibid., 30,243-246.

Akedo, H., and Christensen, H. N. (1962). Transfer of amino-acidsacross the intestine: a new model amino-acid. J. biol. Chem.,237, 113-1 17.

Alexander, H. L., Shirley, K., and Allen, D. (1936). The route ofingested egg white to the systemic circulation. J. clin. Invest., 15,163-167.

Andrews, J. C., Johnston, C. G., and Andrews, K. C. (1936). Theabsorption of cystine, methionine, and cysteic acid fromintestinal loops of dogs. Amer. J. Physiol., 115, 188-193.

Berliner, R. W. (1959). Membrane transport. Rev. mod. Phys., 31,342-348.

Bessman, S. P., Magnes, J., Schwerin, P., and Waelsch, H. (1948).The absorption of glutamic acid and glutamine. J. biol. Chem.,175, 817-823.

Block, K. (1946). The metabolism of l(+)-arginine in the rat. Ibid.,165, 469-476.

Bolton, C., and Wright, G. P. (1937). The absorption of amino-acidsand their distribution in the body fluids. J. Physiol. (Lond.), 89,269-286.

Booth, C. C., and Kanaghinis, T. (1963). A dual mechanism ofabsorption of L-tryptophan. Ibid., 167, 18P.

Bragden, J. H. (1958). On the composition of chyle chylomicrons.J. Lab. clin. Med., 52, 564-570.

Brambell, F. W. R., Halliday, R., and Morris, I. G. (1958). Inter-ference by human and bovine serum and serum proteinfractions with the absorption of antibodies by suckling ratsand mice. Proc. roy. Soc., B, 149, 1-11.

Buglia, G. (1912). Untersuchungen uber die biologische Bedeutungund den Metabolismus der Eiweisstoffe. X. Gesamtstickstoffund Aminosaurestickstoffe im Harn der per os mit Fleischoder auf intravenosem Wege mit den Verdauungsbroduktendes Fleisches ernahrten Tiere. Z. Biol., 58, 162-184.

Butterworth, C. E., Jr., Perez-Santiago, E., Martinez de Jesus, J., andSantini, R. (1959). Studies on the oral and parenteral admini-stration of D(+)-xylose. New Engi. J. Med., 261, 157-164.

Busch, H., Davis, J. R., Honig, G. R., Anderson, D. C., Nair, P. V.,and Nyhan, W. L. (1959). The uptake of a variety of amino-acids into nuclear proteins of tumors and other tissues. CancerRes., 19, 1030-1039.

Cathcart, E. P., and Leathes, J. B. (1906). On the absorption ofproteids from the intestine. J. Physiol. (Lond.), 33, 462-475.

Chase, B. W., and Lewis, H. B. (1934). Comparative studies of themetabolism of amino acids. VI. The rate of absorption ofleucine, valine, and their isomers from the gastrointestinaltract of the white rat. J. biol. Chem., 106, 315-321.

Christensen, H. N. (1960). Reactive sites and biological transport.Advanc. Protein., Chem., 15, 239-314.

- (1962). Intestinal absorption with special reference to amino-acids. Fed. Proc., 21, 37-42.

, Decker, D. G., Lynch, E. L., MacKenzie, T. M., and Powers,J. H. (1947). The conjugated, non-protein, amino-acids ofplasma. V. A study of the clinical significance of peptidemia.J. clin. Invest., 26, 853-859.and Lynch, E. L. (1946). The conjugated, non-protein, amino-acids of plasma. I. Postabsorptive concentrations of humanplasma, serum, and erythrocytes. J. biol. Chem., 163, 741-751.

, and Oxender, D. L. (1960). Transport of amino acids into andacross cells. Amer. J. clin. Nutr., 8, 131-136.

, Parker, H. M., and Riggs, T. R. (1958). Nonexchange ofcarboxyl oxygen in mammalian amino acid transport. J. biol.Chem., 233, 1485-1487.

, Riggs, T. R., and Coyne, B. A. (1954). Effects of pyridoxal andindoleacetate on cell uptake of amino acids and potassium.Ibid., 209, 413-427.

Clark, S. L., Jr. (1959). The ingestion of proteins and colloidalmaterials by columnar absorptive cells of the small intestine insuckling rats and mice. J. biophys. biochem. Cytol., 5, 41-49.

Clarke, E. W., Gibson, Q. H., Smyth, D. H., and Wiseman, G. (1951).Selective absorption of amino-acids from Thiry-Vella loops.J. Physiol. (Lond.), 112, 46P.

Cohnheim, 0. (1901). Die Umwandlung des Eiweiss durch dieDarmwand. Zeit. physiol. Chem., 33, 451-465.

Comline, R. S., Roberts, H. E., and Titchen, D. A. (1951). Histo-logical changes in the epithelium of the small intestine duringprotein absorption in the new-born animal. Nature (Lond.), 168,84-85.

Cordero N., and Wilson, T. H. (1961). Comparison of transportcapacity of small and large intestine. Gastroenterology, 41,500-504.

Cori, C. F. (1926). The absorption of glycine and d, 1-alanine. Proc.Soc. exp. Biol. (N. Y.), 24, 125-126.

Crampton, R. F. and Matthews, D. M. (1964). Kinetics of drugabsorption: methods and interpretation. In Absorption andDistribution of Drugs, pp. 49-63 Ed. Binn's. T. B. Livingstone,Edinburgh and London.

Crane, C. W. (1961). Some aspects of protein digestion and absorptionin health and disease. Postgrad. med. J., 37, 745-754.

-, and Neuberger, A. (1960). The digestion and absorption ofprotein by normal man. Biochem. J., 74, 313-323.

Crane, R. K. (1960). Intestinal absorption of sugars. Physiol. Rev., 40,789-825.and Mandelstam, P. (1960). The active transport of sugars byvarious preparations of hamster intestine. Biochim. biophys.Acta (Amst.), 45, 460-476.

-, and Wilson, T. H. (1958). In vitro method for the study of therate of intestinal absorption of sugars. J. appl. Physiol., 12,145-146.

Csaky, T. Z. (1961). Significance of sodium ions in active intestinaltransport of nonelectrolytes. Amer J. Physiol., 210, 999-1001.

Cummins, A. J., and Almy, T. P. (1953). Studies on the relationshipbetween motility and absorption in the human small intestine.Gastroenterology, 23, 179-190.

Danforth, E., Jr., and Moore, R. 0. (1959). Intestinal absorption ofinsulin in the rat. Endocrinology, 65, 118-123.

Darlington, W. A., and Quastel, J. H. (1953). Absorption of sugarsfrom isolated surviving intestine. Arch. Biochem., 43, 194-207.

Dawson, A. M., and Isselbacher, K. J. (1960). Studies on lipidmetabolism in the small intestine with observations on the roleof bile salts. J. clin. Invest., 39, 730-740.and Matthews, D. M. (1961). An effect of taurocholate onamino-acid production by small intestinal in vitro. J. Physiol.(Lond.), 159, 57-58P.

Draper, H. H. (1958). The absorption of radiolysine by the chick asaffected by penicillin administration. J. Nutr., 64, 33-42.

Dent, C. E., and Schilling, J. A. (1949). Studies on the absorption ofproteins: the amino-acid pattern in the portal blood. (Adden-dum. Christensen, H. N., Conjugated amino-acids in portalplasma of dogs after protein feeding.) Biochem. J., 44, 318-335.

Denton, A. E., and Elvehjem, C. A. (1954). Availability of aminoacids in vivo. J. biol. Chem., 206, 449-454.

, Gershoff, S. N., and Elvehjem, C. A. (1953). A new methodfor cannulating the portal vein of dogs. Ibid., 204,731-735.

Elsden, S. R., Gibson, Q. H., and Wiseman, G. (1950). Selectiveabsorption of amino-acids from the small intestine of the rat.J. Physiol., III, 56P.

Erf, L. A., and Rhoads, C. P. (1940). The glycine tolerance test insprue and pernicious anemia. J. clin. Invest., 19, 409-421.

Feh6r, I., Kertai, P., and Gati, T. (1956). The role of the ATPcontent of the intestinal muocsa in the absorption of glucose.Acta physiol. Acad. Sci. hung., 10, 19-32.

Finch, L. R., and Hird, F. J. R. (1960a). The uptake ofamino acids byisolated segments of rat intestine. I. A survey of factorsaffecting the measurement of uptake. Biochim. biophys. Acta(Amst.), 43, 268-277.- (1960b). The uptake of amino acids by isolated segments ofrat intestine. II. A survey of affinity for uptake from rates ofuptake and competition for uptake. Ibid., 43, 278-287.

Fisher, R. B. (1954). Protein Metabolism. Methuen. London.-, and Parsons, D. S. (1949). A preparation of surviving rat small

intestine for the study of absorption. J. Physiol. (Lond.), 110,36-46.

Folin, 0., and Denis, W. (1912). Protein metabolism from the stand-point of blood and tissue analysis. J. biol. Chem., 11, 87-95.

Fridhandler, L., and Quastel, J. H. (1955). Absorption of amino acidsfrom isolated surviving intestine. Arch. Biochem. 56, 424-440.

Friedberg, F., Tarver, H., and Greenberg, D. M. (1948). The distri-bution pattern of sulfur-labeled methionine in the protein andthe free amino acid fraction of tissues after intravenousadministration. J. biol. Chem., 173, 355-361.


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Fullerton, P. M., and Parsons, D. S. (1956). Absorption of sugars andwater from rat intestine in vivo. Quart. J. exp. Physiol., 41,387-397.

Gibson, Q. H., and Wiseman, G. (1951). Selective absorption ofstereo-isomers of amino acids from loops of the small intestineof the rat. Biochem. J., 48, 426-429.

Gruskay, F. L., and Cooke, R. E. (1955). The gastrointestinal absorp-tion of unaltered protein in normal infants and in infantsrecovering from diarrhea. Pediatrics, 16, 763-769.

Hagihira, H., Ogata, M., Takedatsu, N., and Suda, M. (1960).Intestinal absorption of amino acids. III. Interference betweenamino acids during intestinal absorption. J. Biochem. (Tokyo),47, 139-143.Lin, E. C. C., Samiy, A. H., and Wilson, T. H. (1961). Activetransport of lysine, ornithine, arginine and cystine by theintestine. Biochem. biophys. Res. Commun., 4, 478-481.Wilson, T. H., and Lin, E. C. C. (1962). Intestinal transport ofcertain N-substituted amino acids. Amer. J. Physiol., 203,637-640.

Halliday, R. (1958). The increase in alkaline phosphatase activity ofthe duodenum and decrease in absorption of antibodies by thegut induced in young rats by desoxycorticosterone acetate.J. Physiol. (Lond.), 140, 44-45P.

Harrison, H. E., and Harrison, H. C. (1963). Sodium, potassium, andintestinal transport of glucose, 1-tyrosine, phosphate, andcalcium. Amer. J. Physiol., 205, 107-111.

Heidenhain, R. (1894), Neue Versuche uber die Aufsaugung Dundarm.Pflugers Arch. ges. Physiol., 56, 579-631.

Henderson, Y., and Dean, A. L. (1903). On the question of proteidsynthesis in the animal body. Amer. J. Physiol., 9, 386-390.

Hendrix, B. M., and Sweet, J. E. (1917). A study of amino nitrogenand glucose in lymph and blood before and after the injectionof nutrient solutions in the intestine. J. biol. Chem., 32, 299-307.

Hetenyi, G., Jr., and Winter, M. (1952). Contribution to the mechan-ism of the intestinal absorption of amino acids. Acta. physiol.Acad. Sci., hung., 3, 49-58.

Hill, K. J. (1956). Gastric development and antibody transference inthe lamb, with some observations on the rat and guinea-pig.Quart. J. exp. Physiol., 41, 421-432.

, and Hardy, W. S. (1956). Histological and histochemical observ-ations on the intestinal cells of lambs and kids absorbingcolostrum. Nature (Lond.), 178, 1353-1354.

Hober, R., and Hober, J. (1937). Experiments on the absorption oforganic solutes in the small intestine of rats. J. cell. comp.Physiol., 10, 401-422.

Horvath, I., and Wix, G. (1951). Hormonal influences on glucoseresorption from the intestines. I. Methodical principles, dailyvariations in the absorption of sugar. The proportion betweenthe absorption of glucose and xylose. Acta physiol. Acad. Sci.,hung., 2, 435-443.

Howe, P. E. (1921). An effect of the ingestion of colostrum upon thecomposition of the blood of new-born calves. J. biol. Chem., 49,115-118.

Jacobs, F. A., Coen, L. J., and Hillman, R. S. L. (1960). Influence ofpyridoxine, pyridoxal phosphate, deoxypyridoxine, and 2,4-dinitrophenol on methionine absorption. Ibid., 235, 1372-1375.and Hillman, R. S. L. (1958). Intestinal absorption of themethionine isomers as affected by deoxypyridoxine. Ibid., 232,445-451.

Jervis, E. L., and Smyth, D. H. (1959a). The effect of concentrationsof amino acids on their rate of absorption from the intestine.J. Physiol. (Lond.), 149, 433-441.

(1959b). Competition between enantiomorphs of aminoacids during intestinal absorption. Ibid., 145, 57-65.

(1960). The active transfer of D-methionine by the ratintestine in vitro. Ibid., 151, 51-58.

Johnston, J. M., and Wiggans, D. S. (1958). The absorption in vitroof alanyl-phenylalanine. Biochim. biophys. Acta (Amst.), 27,224-225.

Kekwick, R. A. (1959). The serum proteins of the fetus and young ofsome mammals. In Advanc. Protein Chem., p. 231-254.

Kershaw, T. G., Neame, K. D., and Wiseman, G. (1960). The effectof semistarvation on absorption by the rat small intestine. invitro and in vivo. J. Physiol. (Lond.), 152, 182-190.

Kertai, P., Feher, I., and Gati, T. (1956). The role of the ATP-content of the intestinal mucosa in the absorption of varioussubstances. Acta physiol. Acad. Sci. hung., 10, 33-41.

Kratzer, F. H. (1944). Amino acid absorption and utilization in thechick. J. biol. Chem., 153, 237-247.

Kuroda, Y., and Gimbel, N. S. (1954). Selective disappearance ofstereoisomers of amino acids from the human small intestine.J. appl. Physiol., 7, 148-150.

Kutscher, Fr., and Seemann, J. (1901-1902). Zur Kenntniss derVerdauungsvorgange im Dunndarm. I. Hoppe-Seylers Z.physiol. Chem., 34, 528-543.

Laster, L. and Ingelfinger, F. J. (1961). Intestinal absorption-aspectsof structure, function and disease of the small-intestine mucosa.New Engl. J. Med., 264, 1138-1148; 1192-1200; 1246-1253.

Lathe, G. H. (1943). The effect of the concentration of differentsubstances in the intestinal contents on their absorption fromthe small intestine. Rev. canad. Biol., 2, 134-142.

Lin, E. C. C., and Wilson, T. H. (1960). Transport of L-tyrosine by thesmall intestine in vitro. Amer. J. Physiol., 199, 127-130.Hagihira, H., and Wilson, T. H. (1962). Specificity of thetransport system for neutral amino acids in the hamsterintestine. Ibid., 202, 919-925.

Loewi, 0. (1902). Ueber Eiweisssynthese im Thierkorper. Naunyn-Schmiedeberg's Arch. exp. Path. Pharmak., 48, 303-330.

London, E. S. (1935). Angiostomie und Organestoffwechsel. All-Union-Institut-Exper. Med., Moscow.

, and Kotschneff, N. P. (1934). In welcher Form wird Nahrungsei-weiss resorbiert? I. Mitteilung. Hoppe-Seylers Z. physiol. Chem.,228, 235-242.

Longenecker, J. B., and Hause, N. L. (1959). Relationship betweenplasma amino acids and composition of the ingested protein.Arch. Biochem. Biophys., 84, 46-59.

Matthews, D. M. and Laster, L. (1965a). The kinetics of intestinalactive transport of five neutral amino acids. Amer. J. Physiol.,208, 593-600.and Laster, L. (1965b). Competition for intestinal transportamong five neutral amino acids. Ibid., 601-606.

-, and Smyth, D. H. (1952). Stereochemically specific absorption ofalanine from the intestine into the blood-stream. J. Physiol.,(Lond.), 116, 20-21P.

-, (1954). The intestinal absorption of amino-acid enantio-morphs. Ibid., 126, 96-100.

and Wiseman, G. (1953). Transamination by the small intestineof the rat. Ibid., 120, 55P.

McCance, R. A., Hutchinson, A. 0., Dean, R. F. A., and Jones,P. E. H. (1949). The cholinesterase activity of the serum onnewborn animals and of colostrum. Biochem. J., 45, 493-496.

, and Widdowson, E. M. (1959). The effect of colostrum on thecomposition and volume of the plasma of newborn piglets.J. Physiol. (Lond.), 145, 547-550.

Milne, M. D. (1964). Disorders of amino-acid transport. Brit. med. J.,1, 327-336.

Murlin, J. R., Tomboulian, R. L., and Pierce, H. B. (1937). Absorp-tion of insulin from Thiry-Vella loops of intestine in normaland depancreatized dogs. Amer. J. Physiol., 120, 733-743.

Nasset, R. S. (1957). Role of the digestive tract in the utilization ofprotein and amino acids. J. Amer. med. Ass., 164, 172-177.

Nathans, D., Tapley, D. F., and Ross, J. E. (1960). Intestinal transportof amino acids studied in vitro with L-(131-I) monoiodotyro-sine. Biochim. biophys. Acta (Amst.), 41, 271-282.

Neame, K. D., and Wiseman, G. (1957). The transamination ofglutamic and aspartic acids during absorption by the smallintestine of the dog in vivo. J. Physiol. (Lond.), 135, 442-450.- (1958). The alanine and oxo acid concentrations in mesen-teric blood during the absorption of L-glutamic acid by thesmall intestine of the dog, cat and rabbit in vivo. Ibid., 140,148-155.

Neil, M. W. (1959). The absorption of cystine and cysteine from rat

small intestine. Biochim. J., 71, 118-124.Newey, H., and Smyth, D. H. (1959). The intestinal absorption of

some dipeptides. J. Physiol. (Lond.), 145, 48-56.(1960). Intracellular hydrolysis of dipeptides during

intestinal absorption. Ibid., 152, 367-380.- (1962). Cellular mechanisms in intestinal transfer of aminoacids. Ibid., 164, 527-551.- (1963). Specificity of carriers in intestinal transfer of glycine.Ibid., 165, 74-75P.- (1964). Effects of sugars on intestinal transfer of amino-acids. Nature (Lond.), 202, 400-401.

Paine, C. M., Newman, H. J., and Taylor, M. W. (1959). Intestinalabsorption of methionine and histidine by the chicken. Amer.J. Physiol., 197, 9-12.


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426 D. M. Matthews and Leonard Laster

Parkinson, T. M., and Olson, J. A. (1963). Inhibitory effects of bileacids on the uptake, metabolism and transport of water-soluble substances in the small intestine of the rat. Life Sci., 6,393-398.

Parshin, A. N., and Rubel, L. N. (1951). The nature of products ofprotein digestion that are absorbed from the intestine into theblood. (In Russian.) Dokl. Akad. Nauk. SSSR., 7, 313-315(abstract in Chem. Abstr., 45, 7206).

Parsons, B. J., Smyth, D. H., and Taylor, C. B. (1958). The action ofphlorrhizin on the intestinal transfer of glucose and waterin vitro. J. Physiol. (Lond.), 144, 387-402.

Pinsky, J., and Geiger, E. (1952). Intestinal absorption of histidine asinfluenced by tryptophan in the rat. Proc. Soc. exp. Biol. (N. Y.),81, 55-57.

Quastel, J. H. (1961). Technics for studies of intestinal absorptionin vitro. Meth. med. Res., 9, 273-286.

Ratner, B., and Gruehl, H. L. (1934). Passage of native proteinsthrough the normal gastro-intestinal wall. J. clin. Invest., 13,517-532.

Reynolds, E. H., Hallpike, J. F., Phillips, B.M., and Matthews, D. M.(1965). Reversible absorptive defects in anticonvulsant meg-aloblastic anaemia. J. clin. Path., in the press.

Rosenberg, T. (1954). The concept and definition of active transport.Symp. Soc. exp. Biol., 8, 27-41.

Schofield, F. A., and Lewis, H. B. (1947). A comparative study of themetabolism of a-alanine, 3-alanine, serine, and isoserine. I.Absorption from the gastrointestinal tract. J. biol. Chem., 168,439-445.

Schmengler, F. E. (1933). O0ber die Permeabilitat getrockneterKollodiummembranen fur Aminosauren im Vergleich zuorganischen Nichtelectrolyted. Pflugers Arch. ges. Physiol., 232,591-603.

Sheff, M. F., and Smyth, D. H. (1955). An apparatus for the study ofin vivo intestinal absorption in the rat. J. Physiol. (Lond.), 128,67P.

Shishova, 0. A. (1956). Role of phosphorylation in absorption ofamino acids in intestine. Biochemistry, 21, 105-113. (Russian textBiokhimiya, 21, 111-118).(1959). The effect of phosphorylation on the absorption ofvarious amino acids. Ibid., 24, 480-485. (Russian text, Ibid.,24, 514-520.)

Smith, E. L. (1948). The isolation and properties of the immuneproteins of bovine milk and colostrum and their role inimmunity: a review. J. Dairy Sci., 31, 127-138.

Smyth, D. H. (1961a). Intestinal absorption. Proc. roy. Soc. Med.,54, 769-773.(1961b). Methods for study of intestinal absorption in vivo.Meth. med. Res., 9, 260-272.and Taylor, C. B. (1957). Transfer of water and solutes by anin vitro intestinal preparation. J. Physiol. (Lond.), 136, 632-648.and Whaler, B. C. (1953). Apparatus for the in vitro study ofintestinal absorption. Ibid., 121, 2-3P.

Sols, A., and Ponz, F. (1947). A new method for the study of intestinalabsorption. Rev. espan. Fisiol., 3, 207-21 1.

Spencer, R. P., Bow, T. M., and Markulis, M. A. (1962). Aminogroup requirements for in vitro intestinal transport of aminoacids. Amer. J. Physiol., 202, 171-173.

, and Knox, W. E. (1960). Comparative enzyme apparatus of thegut mucosa. Fed. Proc. 19, 886-897.

, and Samiy, A. H. (1960). Intestinal transport of L-tryptophanin vitro: inhibition by high concentrations. Amer. J. Physiol.,199, 1033-1036.- (1961). Intestinal absorption of L-phenylalanine in vitro.Ibid., 200, 501-504.

- and Zamcheck, N. (1961). Presence of glutaminase I in ratintestine. Gastroenterology, 40, 423-426.

Stein, W. H., Bearn, A. G., and Moore, S. (1954). The amino acidcontent of the blood and urine in Wilson's disease. J. clin.Invest., 33, 410-419.

, and Moore, S. (1954). The free amino acids of human bloodplasma. J. biol. Chem., 211, 915-926.

Sugawa, T., Akedo, H., and Suda, M. (1960). Intestinal absorption ofamino acids. II. Isotopic studies on the amino acids absorptionfrom the intestine using S35 DL-methionine and C14 DL-valine. J. Biochem. (Tokyo), 47, 131-138.

Sussman, H., Davidson, A., and Walzer, M. (1928). Absorption ofundigested proteins in human beings. III. The absorption ofunaltered egg protein in adults. Arch. intern. Med., 41, 409-414.

Tapley, D. F., Herz, R., Jr., Ross, J. E., Deuel, T. F., and Leventer, L.(1960). Glucuronide formation in active transport of aminoacid and thyroxine analogues by rat intestine. Biochim.biophys. Acta (Amst.), 43, 344-345.

Ussing, H. H. (1957). General principles and theories of membranetransport. In Metabolic Aspects of Transport across CellMembranes, edited by Q. R. Murphy, pp. 39-56. University ofWisconsin Press, Madison.

Van Slyke, D. D., and Meyer, G. M. (1912). The amino-acid nitrogenof the blood. Preliminary experiments on protein assimilation.J. biol. Chem., 12, 399-410.,- (1913-14). The fate of protein digestion products in the body.

III. The absorption of amino acids. Ibid., 16, 197-212.Verzar, F., and McDougall, E. J. (1936). Absorption from the Intestine.

Longmans, London.Visscher, M. B., Roepke, R. R., and Lifson, N. (1945). Osmotic and

electrolyte concentration relationships during the absorptionofautogenous serum from ileal segments. Amer. J. Physiol., 144,457-463.

Voit, C., and Bauer, J. (1869). Ueber die Aufsaugang im Dick-undDunndarme. Z. Biol., 5, 536-570.

Wang, H. L., and Waisman, H. A. (1961). Phenylalanine tolerancetests in patients with leukemia. J. Lab. clin. Med., 57, 73-77.

Whaler, B. C. (1955). The metabolism of amino acids by the smallintestine. J. Physiol. (Lond.), 130, 278-290.

Wiggans, D. S., and Johnston, J. M. (1959). The absorption ofpeptides. Biochim. biophys. Acta (Amst.), 32, 69-73.

Wilbrandt, W., and Rosenberg, T. (1961). The concept of carriertransport and its corollaries in pharmacology. Pharmacol. Rev.,13, 109-183.

Wilson, S. J., and Walzer, M. (1935). Absorption of undigestedproteins in human beings. IV. Absorption of unaltered eggprotein in infants and in children. Amer. J. dis. Child., 50,49-54.

Wilson, T. H. (1962). Intestinal Absorption, pp. 122-123. Saunders,Philadelphia and London.

(1964). Structure and function of the intestinal absorptive cell.In The Cellular Functions of Membrane Transport, edited byJ. F. Hoffman, pp. 215-229. Prentice Hall, New Jersey.

, and Lin, E. C. C. (1960). Active transport by intestines of fetaland newborn rabbits. Amer. J. Physiol., 199, 1030-1032.

, Landau, B. R., and Jorgensen, C. R. (1960). Intestinaltransport of sugars and amino acids. Fed. Proc., 19, 870-875.and Wiseman, G. (1954). The use of sacs of everted smallintestine for the study of the transference of substances fromthe mucosal to the serosal surface. J. Physiol. (Lond.), 123,116-125.

Wiseman, G. (1951). Active stereochemically selective absorption ofamino acids from rat small intestine. Ibid., 114, 7-8P.

(1953). Absorption of amino-acids using an in vitro technique.Ibid., 120, 63-72.

- (1955). Preferential transference of amino-acids from amino-acidmixtures by sacs of everted small intestine of the goldenhamster. (Mesocricetus auratus). Ibid., 127, 414-422.

- (1956). Active transport of amino acids by sacs of everted smallintestine of the golden hamster (Mesocricetus auratus). Ibid.,133, 626-630.

(1957). Effects of pyridoxal on the active transport of amino acidsby sacs of everted intestine of the golden hamster. (Mesocricetusauratus). Ibid., 136, 203-207.

- (1961). Sac of everted intestine technic for study of intestinalabsorption in vitro. Meth. med. Res., 9, 287-292.

(1964). Absorption from the intestine. Academic Press. Londonand New York.Neame, K. D., and Ghadially, F. N. (1959). Effect of sarcomaRD3 on intestinal active absorption of glucose and L-histidine.Brit. J. Cancer, 13, 282-287.

Wolff, J. (1964). Transport of iodide and other anions in the thyroidgland. Physiol. Rev., 44, 45-90.

Wright, R. D., and Wynn, B. (1951). The digestion of protein in thealimentary canal. Austr. J. exp. Biol. med. Sci., 29, 281-284.

Yuile, C. L., O'Dea, A. E., Lucas, F. V., and Whipple, G. H. (1952).Plasma protein labeled with lysine-E-C14: its oral feeding andrelated protein metabolism in the dog. J. exp. Med., 96,247-254.


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